The Kinetics of Fatty Acid Uptake By Paramecium Tetraurelia

ABSTRACT. The kinetics of radiolabeled fatty acid uptake by the ciliate Paramecium tetraurelia was examined on a homologous series of saturated, straight chain fatty acids of even carbon numbers. Uptake rates increased with chain length from acetate to palmitate. Saturation kinetics was demonstrated for most fatty acids examined, thus ruling out simple diffusion as the major mechanism for fatty acid transport and implicating carrier-mediated, facilitated transport as the major mechanism. Data from most competitive inhibition experiments were too scattered to determine the number of transporter systems present. Cholesterol uptake also exhibited saturation kinetics and hence other sterols, which can satisfy this nutritional requirement, may also be transported by a carrier-mediated mechanism. the uptake of the essential fatty acid oleate was faster than those observed for the saturated acids and could not be explained by only one transport mechanism. Therefore, fatty acid transport also occurs via other kinetically significant routes.     

Physiological Characterization of Root Zn2+ Absorption and Translocation to Shoots in Zn Hyperaccumulator and Nonaccumulator Species of Thlaspi

Radiotracer techniques were employed to characterize 65Zn2+ influx into the root symplasm and translocation to the shoot in Thlaspi caerulescens, a Zn hyperaccumulator, and Thlaspi arvense, a nonaccumulator. A protocol was developed that allowed us to quantify unidirectional 65Zn2+ influx across the root-cell plasma membrane (20 min of radioactive uptake followed by 15 min of desorption in a 100 [mu]M ZnCl2 + 5 mM CaCl2 solution). Concentration-dependent Zn2+ influx in both Thlaspi species yielded nonsaturating kinetic curves that could be resolved into linear and saturable components. The linear kinetic component was shown to be cell-wall-bound Zn2+ remaining in the root after desorption, and the saturable component was due to Zn2+ influx across the root-cell plasma membrane. This saturable component followed Michaelis-Menten kinetics, with similar apparent Michaelis constant values for T. caerulescens and T. arvense (8 and 6 [mu]M, respectively). However, the maximum initial velocity for Zn2+ influx in T. caerulescens root cells was 4.5-fold higher than for T. arvense, indicating that enhanced absorption into the root is one of the mechanisms involved in Zn hyperaccumulation. After 96 h 10-fold more 65Zn was translocated to the shoot of T. caerulescens compared with T. arvense. This indicates that transport sites other than entry into the root symplasm are also stimulated in T. caerulescens. We suggest that although increased root Zn2+ influx is a significant component, transport across the plasma membrane and tonoplast of leaf cells must also be critical sites for Zn hyperaccumulation in T. caerulescens.     

Blood-Brain Barrier Transport of 1-Aminocyclohexanecarboxylic Acid, a Nonmetabolizable Amino Acid for In Vivo Studies of Brain Transport

Regional transport of 1-aminocyclohexanecarboxylic acid (ACHC), a nonmetabolizable amino acid, across the blood-brain barrier was studied in pentobarbital-anesthetized rats using an in situ brain perfusion technique. The concentration dependence of influx was best described by a model with a saturable and a nonsaturable component. Best-fit values for the kinetic constants of the frontal cortex equaled 9.7 X 10(-4) mumol/s/g for Vmax, 0.054 mumol/ml for Km, and 1.0 X 10(-4) ml/s/g for KD in the absence of competing amino acids. Saturable influx could be reduced by greater than 85% by either L-phenylalanine or 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid, consistent with transport by the cerebrovascular neutral amino acid transport system. The transport Km for ACHC was one-fifth that for the more commonly used homologue, 1-aminocyclopentanecarboxylic acid, and was similar to values for several natural amino acids, such as L-methionine, L-isoleucine, and L-tyrosine. The results indicate that ACHC may be a useful probe for in vivo studies of amino acid transport into brain.     

The uptake and metabolism of urea by Chara australis: IV. Symport with sodium—A slip model for the high and low affinity systems

We have previously investigated the electrogenic influx of urea in Chara, and the urea- and sodium-dependent membrane current. We have shown that there is a sodium-stimulated component of urea influx and a urea-stimulated component of sodium influx, and that these are of the same size. We conclude that the electrogenic inward transport of urea, and of its analogues acetamide and acrylamide, is by sodium symport, with a stoichiometric ratio of 1∶1. The kinetics of the fluxes and currents show two different K _M values for sodium in different cells and two different kinds of kinetics for the effect of urea on membrane current, one of which fits the Michaelis-Menten equation, while the other shows a maximum and fits the difference of two Michaelis-Menten terms, suggesting a phenomenon like cis-inhibition. Similarities in kinetic characteristics between the inhibitory site and the electrically silent uptake site (System II) lead us to suggest that the same protein may be responsible for both the low-K _M, electrogenic influx of urea (System I) and the high-K _M, electrically silent influx by System II. We suggest a “slip” model for urea uptake in Chara.     

Transport of amino acids by isolated gills of the mussel Mytilus californianus Conrad.

The unidirectional influx of cycloleucine into in vitro preparations of gill tissue of the mussel, Mytilus californianus, was determined. Influx was found to be linear for at least an hour, and the kinetics of cycloleucine influx conformed to Michaelis-Menten type kinetics. The transport mechanism(s) for cycloleucine is relatively specific for the L-enantiomorph of neutral amino acids, and is capable of accumulating cycloleucine to intracellular concentrations much higher than those of the surrounding medium. Evedence is presented that the transport of amino acids by gill tissue plays a significant role in whole animal nutrition.     

Kinetics of Myo-inositol transport in rat ocular lens

Myo-inositol (MI) influx as a function of concentration in rat lens consisted of a saturable component, fit by a rectangular hyperbola, and a linear component which was more distinct at high myo-inositol concentrations suggesting passive diffusion. The hyperbolic component was half-maximally saturated (K_t) at 61.3 μM and had a maximal transport rate (J_max) of 44.6 μMol/kg wet wt/h. The linear component had an apparent permeability coefficient of 1.44 × 10^?6 s^?1. Sorbitol, which distributed rapidly in the extracellular space (6.83 ml/100 g wet wt), also appeared to enter the intracellular space with a permeability coefficient of 1.37 × 10^?6 s^?1, similar to that of myo-inositol. The influx of myo-inositol was critically dependent on the concentration of extracellular sodium consistent with a sodium-myo-inositol contransport. The kinetics of influx activation by sodium suggested an apparent 2:1 coupling ratio for sodium and myo-inositol. When potassium was used as sodium substitute, a significantly stronger influx inhibition was observed than with nondepolarizing sodium substitutes, indicating that myoinositol was driven by the electrochemicl gradient of sodium rather than the chemical gradient only. Reducing the extracellular Na concentration increased the MI concentration at which transport was half-maximally activated, suggesting an ordered binding sequence of Na followed by MI. Myo-inositol influx was competitively inhibited by phlorizin with an inhibitory coefficient (K_i) of 35 μM. Phloretin also was capable of inhibition but with a much lesser efficacy. Myoinositol desaturates from the lens at a rate of 0.00862 h^?1. Approximately 19% of the efflux can be inhibited with phlorizin, suggesting that it represents carrier-mediated flux. The phlorizin insensitive flux has a rate of 0.00695 h^?1 or 1.93 × 10^?6 s^?1, similar to the Na-independent passive influx. MI influx is due to a Na-dependent, phlorizin-sensitive active transport while the efflux consists largely of a phlorizin-independent passive leakage. © 1995 Wiley-Liss, Inc.     

NUTRIENT TRANSPORT BY THE CRUSTACEAN GASTROINTESTINAL TRACT: RECENT ADVANCES WITH VESICLE TECHNIQUES

1. Techniques are described for producing purified brush-border membrane vesicles (BBMV) of crustacean hepatopancreas which can be used to examine the characteristics of solute transport at the apical pole of hepatopancreatic epithelial cells. 2. Hepatopancreatic BBMV illustrated Na-dependent, carrier-mediated sugar transport which was electrogenic and sensitive to pH. Increased proton concentration lowered the Michaelis-Menten constant for glucose transport and increased the apparent diffusional permeability of the membrane to sugar. 3. Transports of L-alanine and L-lysine by hepatopancreatic BBMV were Na-independent, carrier-mediated, and strongly sensitive to transmembrane electrical potential after protonation at acidic pH. L-alanine and L-lysine were competitive inhibitors of each other for influx into BBMV and also illustrated trans-stimulation, suggesting that both amino acids use the same transfer mechanism. L-Leucine was a non-competitive inhibitor of L-lysine influx and may employ a distinct Na-independent transport process. 4. L-glutamate transport after protonation at acidic pH was Na-dependent, suggesting that a different transport mechanism was responsible for its movement across hepatopancreatic BBMV than that facilitating the transfer of alanine or lysine. 5. Preliminary experiments indicate the presence of Na/H antiport in hepatopancreatic BBMV, providing, for the first time, a possible mechanism for gastrointestinal luminal acidification in crustaceans. 6. A proposed model for nutrient transport by crustacean hepatopancreatic BBMV is presented which suggests that transapical transfers of both sugars and amino acids are strongly influenced by in vivo luminal acidification. Luminal protons have at least two major effects on nutrient transport in these animals: (a) titration of sugar transport proteins with subsequent stimulatory effects on influx kinetic constants; (b) protonation of luminal amino-acid-charged moieties and conversion into appropriate substrates for transport by either Na-dependent or Na-independent, membrane-potential-sensitive carrier proteins.     

Kinetics and Block of Dopamine Uptake in Synaptosomes from Rat Caudate Nucleus

The dopamine (DA) uptake system in mammalian nerve terminals was studied by measuring the unidirectional influx of tritiated DA into synaptosomes prepared from rat caudate nucleus. Two distinct time-dependent components of DA uptake were observed. The principal component was saturable with respect to DA concentration, required both external Na and Cl, and was competitively blocked by micromolar concentrations of the psychotropic agents cocaine, benztropine, nomifensine, amphetamine, and methamphetamine. This principal component of uptake has the properties expected for a carrier-mediated transport system. The second component, which accounted for about 10-30% of the DA uptake at 2 microM DA, was not saturable, and was independent of external Na, Cl, and blockers of the carrier-mediated system. The saturable, Na-dependent component had an apparent Km(DA) of about 0.5 microM. The dependence of DA uptake on external Na was sigmoid [Hill coefficient = 2; Ka(Na) = 45 mM] whereas the dependence on Cl was best described by a rectangular hyperbola [Ka(Cl) = 15 mM]. Depolarizing conditions (elevated external K) reduced the rate of DA influx. The data are consistent with a carrier-mediated DA transport mechanism in which each DA molecule entering the nerve terminal via the carrier is accompanied by two or more Na ions and one Cl ion in a rheogenic process carrying one or more net positive charges into the cell. Net, concentrative accumulation of DA inside nerve terminals may be accomplished by utilizing the Na electrochemical gradient to drive DA against its electrochemical gradient via this carrier system.     

Sucrose uptake by developing soybean cotyledons

Sucrose uptake by excised developing soybean cotyledons shows a biphasic dependence on sucrose concentration. At concentrations less than about 50 millimolar external sucrose, uptake can be described as a carrier-mediated process, with a K_m of 8 millimolar. At higher external sucrose concentrations, a linear dependence becomes apparent, which suggests the participation of a nonsaturable component in total uptake. Sucrose absorption is dependent on the presence of an electrochemical potential gradient for protons since agents interfering with the generation or maintenance of this gradient (NaN₃ or carbonylcyanide-m-chlorophenyl hydrazone) decrease sucrose transport to a level at or below that predicted from the operation of the noncarrier-mediated process alone. The saturable component of sucrose uptake is also sensitive to the sulfhydryl-modifying compounds N-ethylmaleimide and p-chloro-mercuribenzenesulfonate. The thiol-reducing agent diethioerythritol reverses fully the p-chloro-mercuri-benzenesulfonate inhibition, but not that of N-ethyl maleim de. Sucrose transport is sensitive to external pH, being decreased at high pH₀. Since sucrose-induced depolarization of the membrane potential and carrier-mediated sucrose influx show similar pH-dependence, inhibitor sensitivity, and values of K_m for sucrose, a sucrose/proton contransport process appears to operate in developing soybean cotyledon cells. Measurement of free space and intracellular sucrose concentrations in vivo suggests that the carrier-mediated process is fully saturated and that sucrose transport may be limiting for sucrose accumulation by the developing seed.     

Auxin Transport in Suspension-Cultured Soybean Root Cells : II. Anion Effects on Carrier-Mediated Uptake

To test the hypothesis that the carrier-mediated component of the indoleacetic acid (IAA) influx involves an electrogenic proton/IAA anion symport, the effects on the IAA influx of salts expected to depolarize the membrane potential were examined in suspension-cultured soybean (Glycine max [L.] Merr.) root cells. Although KCl does inhibit carrier-mediated uptake, the effect is specific to the anion at low concentrations and not due to more general processes such as changes in ionic or osmotic strength. Other anions such as bromide, iodide, and fluoride inhibit the carrier more strongly. Because potassium iminodiacetate, which is also expected to depolarize the membrane potential, has no inhibitory effect on the IAA influx, there is no evidence for the involvement of the membrane potential in carrier-mediated uptake. It is therefore most likely that in soybean cells, if carrier-mediated uptake occurs via a proton symport, the H⁺:IAA— stoichiometry is 1:1. At concentrations greater than 70 millimolar, sorbitol, a nonionic osmoticum, inhibits carrier-mediated IAA uptake. The effects of specific anions and osmotic potential on the uptake carrier necessitates the reevaluation of other auxin transport studies in which KCl was routinely used as an agent with which to depolarize the membrane potential.     

The Role of Iron-Deficiency Stress Responses in Stimulating Heavy-Metal Transport in Plants

Plant accumulation of Fe and other metals can be enhanced under Fe deficiency. We investigated the influence of Fe status on heavy-metal and divalent-cation uptake in roots of pea (Pisum sativum L. cv Sparkle) seedlings using Cd²⁺ uptake as a model system. Radiotracer techniques were used to quantify unidirectional ¹⁰⁹Cd influx into roots of Fe-deficient and Fe-sufficient pea seedlings. The concentration-dependent kinetics for ¹⁰⁹Cd influx were graphically complex and nonsaturating but could be resolved into a linear component and a saturable component exhibiting Michaelis-Menten kinetics. We demonstrated that the linear component was apoplastically bound Cd²⁺ remaining in the root cell wall after desorption, whereas the saturable component was transporter-mediated Cd²⁺ influx across the root-cell plasma membrane. The Cd²⁺ transport system in roots of both Fe-deficient and Fe-sufficient seedlings exhibited similar Michaelis constant values, 1.5 and 0.6 μm, respectively, for saturable Cd²⁺ influx, whereas the maximum initial velocity for Cd²⁺ uptake in Fe-deficient seedlings was nearly 7-fold higher than that in Fe-grown seedlings. Investigations into the mechanistic basis for this response demonstrated that Fe-deficiency-induced stimulation of the plasma membrane H⁺-ATPase did not play a role in the enhanced Cd²⁺ uptake. Expression studies with the Fe²⁺ transporter cloned from Arabidopsis, IRT1, indicated that Fe deficiency induced the expression of this transporter, which might facilitate the transport of heavy-metal divalent cations such as Cd²⁺ and Zn²⁺, in addition to Fe²⁺.     

A Reanalysis of the Two-Component Phloem Loading System in Beta vulgaris

Kinetic analysis of [¹⁴C]sucrose loading into sugar beet leaf discs revealed the presence of two transport components. At low exogenous sucrose concentrations, a saturable component, which exhibited Michaelis-Menten characteristics, was the main mode of transport. At concentrations greater than 50 millimolar, phloem loading was dominated by a linear component which appeared to operate as a first order kinetic transport process. Over the exogenous sucrose concentrations employed, influx could be described by the equation v = V_maxS/(S + K_m) + kS. Influx via both processes was strongly pH-dependent. Evidence is presented that the linear component was not explicable in terms of simple diffusion, or exchange diffusion, into either mesophyll or minor vein phloem tissue. Extensive metabolic conversion of sucrose was not a factor contributing to influx at high external sucrose concentrations. At present, it is believed that both components operate in parallel at the membrane bounding the sieve element-companion cell complex. The saturable component is identified with sucrose-H⁺ cotransport. While the significance of the linear component has been established, its nature remains to be elucidated.     

Sodium and Sugar Fluxes across the Mucosal Border of Rabbit Ileum

Unidirectional influxes of sugars and Na from muscosal solution into the cells of rabbit ileum have been examined. The influxes of glucose, galactose, and 3-0-methyl glucose (3 MG) follow Michaelis-Menten type kinetics and are markedly dependent on the presence ofNa in the mucosal solution. For 3 MG, reduction of Na concentration causes a decrease in maximal rate of influx and little change in the "apparent Michaelis constant." There appeared to be little mediated entry of 3 MG into the cells from Na-free solution. The influx of Na was increased by the presence of 3 MG in the mucosal solution and at all Na concentrations tested, there was a 1:1 ratio between sugar influx and the sugar-dependent Na influx. On the basis of these observations, a model has been developed for the sugar transport system involving a transport site that combines with both sugar and Na.     

Adenosine and tubercidin binding and transport in Chinese hamster ovary and Novikoff rat hepatoma cells

The uptake of adenosine and tubercidin by control and ATP-deleted wild-type and adenosine kinase-deficient cells was measured by rapid kinetic techniques. Adenosine deamination was inhibited by pretreatment with 2-deoxy-coformycin. Control wild-type cells phosphorylated adenosine so rapidly that the kinetics of transport per se could not be assessed unambiguously. ATP depletion and adenosine kinase deficiency did not abolish the conversion of adenosine to nucleotides, but reduced it to such an extent that initial velocities of uptake could be safely construed as transport velocities in both zerotrans and equilibrium exchange modes. The same was true for tubercidin, which was not phosphorylated in adenosine kinase-deficient cells. It accumulated intracellularly, however, to concentrations 50 to 120% higher than those in the extracellular space, apparently due to binding to some intracellular component(s). Binding was not saturated up to a concentration of 200 μM, but seemed to be slow relative to transport. Fits of appropriate integrated rate equations based on the simple carrier model to uptake time courses obtained under these conditions yielded Michaelis-Menten constants for adenosine and tubercidin transport of 100 to 200 μM and maximum velocities of 10 to 30 pmol/μl cell H₂O ? sec, whereas the rate of intracellular phosphorylation was maximal at concentrations between 2 and 8 μM. The first-order rate constant (V_max/K_m) for adenosine phosphorylation, however, seemed to be appreciably higher than that for its transport. This indicates that at physiological concentrations, which fall in the first-order range for both processes, adenosine trapping is very efficient. Adenosine, tubercidin, tricyclic nucleoside, 2′-deoxyadenosine, and 3′-deoxyadenosine all inhibited uridine and thymidine transport to about the same extent, whereas pyrazofurin was signficantly less effective.     

Mechanisms for the facilitated diffusion of substrates across cell membranes

Two classes of theoretical mechanisms for protein-mediated, passive, transmembrane substrate transport (facilitated diffusion) are compared. The simple carrier describes a carrier protein that exposes substrate influx and efflux sites alternately but never both sites simultaneously. Two-site models for substrate transport describe carrier proteins containing influx and efflux sites simultaneously. Velocity equations describing transport by these mechanisms are derived. These equations take the same general form, being characterized by five experimental constants. Simple carrier-mediated transport is restricted to hyperbolic kinetics under all conditions. Two-site carrier-mediated transport may deviate from hyperbolic kinetics only under equilibrium exchange conditions. When both simple- and two-site carriers display hyperbolic kinetics under equilibrium exchange conditions, these models are indistinguishable by using steady-state transport data alone. Seven sugar transport systems are analyzed. Five of these systems are consistent with both models for sugar transport. Uridine, leucine, and cAMP transport by human red cells are consistent with both simple- and two-site models for transport. Human erythrocyte sugar transport can be modeled by simple- and two-site carrier mechanisms, allowing for compartmentalization of intracellular sugars. In this instance, resolution of the intrinsic properties of the human red cell sugar carrier at 20 degrees C requires the use of submillisecond transport measurements.     

Markedly altered membrane transport and intracellular binding of vincristine in multidrug-resistant Chinese hamster cells selected for resistance to vinca alkaloids

Studies of a multidrug-resistant variant (DC-3F/VCRd-5L) of Chinese hamster lung cells selected for resistance to vinca alkaloids revealed marked alterations in transport and intracellular binding of [3H]vincristine compared to parental DC-3F cells. Influx of [3H]vincristine in DC-3F cells appears to be an equilibrating, but mediated, process. Although saturation kinetics for [3H]vincristine influx were not demonstrated, an extremely high temperature-dependence (Q10 27-37 degrees C = 5-6) and trans-inhibition of influx following preloading of cells with nonradioactive vincristine argue in favor of a carrier-mediated process. Efflux of [3H]vincristine from parental cells conformed to first-order kinetics (t1/2 37 degrees = 3.6 +/- 0.4) and exhibited a lower temperature-dependence (Q10 27-37 degrees C = 3-3.5) than influx. In variant vs. parental cells, influx of [3H]vincristine was reduced 24-fold and efflux was increased two-fold, accounting for the large (approximately 48-fold) reduction in steady-state level of exchangeable drug accumulating in variant cells. Otherwise, transport of [3H]vincristine in these cells showed characteristics similar to parental DC-3F cells. Also, the rate and amount of intracellular binding of [3H]vincristine in variant cells was almost 40-fold lower than in parental cells. These alterations in influx and efflux of [3H]vincristine and its intracellular binding appear to account, at least to a major extent, for the high level of resistance (2,750-fold) of this variant to vinca alkaloids. In contrast, cross-resistance of this variant to daunomycin (178-fold) could be explained only minimally by a transport alteration. Only a two-fold increase in efflux of [3H]daunomycin was demonstrated in variant vs. parental cells along with some decrease in intracellular binding. Influx of [3H]daunomycin was unaltered. In view of these results, we conclude that these two agents most likely do not share the same route for entry in these cells but might share the same efflux route.     

Transport Interactions between Paraquat and Polyamines in Roots of Intact Maize Seedlings

Interactions between absorption of paraquat and the polyamines putrescine, cadaverine, and spermine in roots of intact maize (Zea mays L. cv 3377 Pioneer) seedlings were examined. Concentration-dependent kinetics for paraquat and putrescine influx were similar and both kinetic curves could be resolved into a linear and a saturable component. The linear component was previously shown to represent cell wall/membrane binding. The saturable components for paraquat and putrescine uptake, which represent influx across the plasmalemma, had K_m values of 98 and 120 micromolar, respectively, and V_max values of 445 and 456 nanomoles per gram fresh weight per hour, respectively. Lineweaver-Burk transformation of the saturable component of paraquat influx in the presence of varying concentrations of putrescine indicated that the diamine competitively inhibited the saturable component of paraquat uptake. Reciprocal experiments similarly demonstrated that paraquat competitively inhibited the saturable component of putrescine uptake. Competitive inhibition of both paraquat and putrescine influx could also be demonstrated with the diamine cadaverine, which has a charge distribution similar to that of paraquat and putrescine. In contrast, the larger, tetravalent polyamine spermine appeared to noncompetitively inhibit the influx of paraquat and putrescine. These results strongly suggest that paraquat enters maize root cells via a carrier system that normally functions in the transport of diamines with a charge distribution similar to that of paraquat.     

The Salt Absorption Isotherm

It is shown that the absorption isotherm of rubidium by excised barley roots can be explained either by two uptake mechanisms following Michaelis-Menten kinetics or by two mechanisms, one actively transporting salts into the tissue (the pump), the other one being more passive in nature (the leak), operating in either direction, depending on external and internal substrate concentration. Kinetic data are thus consistent with more than one transport model. It was further demonstrated for the pair K-Na, that a competitor not only reduces salt uptake but can also reverse the direction of net flux. This observation cannot be explained by classical enzyme kinetics, it is, however, consistent with the pump and leak system. Just as Michaelis-Menten kinetics, the pump and leak system can explain ion competition, in addition it offers a possible explanation of the Viets' effect and it can explain the time curve of absorption.     

The pH dependence of exchange transport of glucose in human erythrocytes

In glucose exchange transport into red blood cells the rate of glucose uptake showed two pH dependent maxima, with the larger at approximately pH 7.5 and the smaller one at pH 3. In the studied pH range the relation between the rate of glucose uptake and the substrate concentration followed Michaelis-Menten kinetics. While the maximal velocity (V) reflected the pH changes of the media, the Michaelis constant (K_m) remained constant. The dissociation constants of the groups of the free carrier and the carrier-glucose complex were the same. The pK of the acidic group was 5.2 and of the basic group 9.5. Glucose was not bound to groups of the carrier which dissociated protons in the pH range of three to nine. By rearranging the equation for the pH dependence of the relative influx a more definitive graphic determination of the pK values was produced.     

Glucose Transport in the Malarial (Plasmodium lophurae) Infected Erythrocyte*

Plasmodium lophurae-infected red blood cells utilized considerably greater quantities of glucose than did uninfected duckling red cells. Kinetic analysis of glucose transport showed: (A). Below a concentration of 2 mM in the medium the uptake process followed Michaelis-Menten kinetics (carrier-mediated facilitated diffusion) whereas at concentrations greater than this simple diffusion became the main mode of entry. (B). The apparent transport constants, K_t, for normal and infected cells were similar. However there was an 8-fold increase in the maximal velocity, V_max, for infected cells. (C). “Free” malaria parasites had a significantly lower K_t and a higher V_max than did normal or infected red cells. Entry and exit studies with the nonmetabolizable sugar analog, 3-0-methyl glucose, demonstrated that the enhanced rate of uptake by infected cells involved an increase in the simple diffusion component and the degree of enhancement was correlated with the size of the intracellular parasite. Competition experiments suggested that in the malaria-infected cell one locus is involved in the carrier-mediated transport of glucose, mannose and galactose whereas another locus transports fructose and/or glycerol. These results indicate that the enhanced entry of glucose into the malaria-infected red cell is a consequence of factors other than increased glucose catabolism by the host-parasite complex, and the host cell's capacity to take up greater quantities of sugar directly involves the growing intracellular plasmodium.